日本細胞生物学会Japan Society for Cell Biology

Immunofluorescence by Zenon

AuthorAkihiro Harada
AffiliationDepartment of Cell Biology, Graduate School of Medicine, Osaka University
Contactinquiry about this protocol
Home Pagehttp://www.med.osaka-u.ac.jp/pub/acb/eng.html
Keyword
Published2015-08-26Last Update2015-08-26
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Summary

Methods

Detail  *Click photo to see a large size.

    • Wash the sections with PBS once
    • Block the nonspecific staining by treating the sections in 5%Normal donkey Serum/PBS
    • Treat the sections with diluted primary antibody in 5%Normal donkey Serum/PBS, 37C 1hr
    • Wash the sections with PBS several times
    • Block the nonspecific staining by treating the sections in 5%Normal donkey Serum/PBS
    • Treat the sections with diluted secondary antibodies in 5%Normal donkey Serum/PBS, 37C 30min
    • Wash the sections with PBS several times
    • Block the nonspecific staining by treating the sections in 5%Normal Goat Serum/PBT(0.1%TritonX-100/PBS)
    • Treat the sections with diluted tertiary antibodies (labeled by Zenon) in PBT, RT 1hr
    • Wash the sections with PBS several times
    • Postfix the sections with 4%PFA/PBS,RT 15min
    • Wash the sections with PBS once
    • Embedding
    • Protocol for making tertiary antibody

      1) Mix antibody solution 10μℓ+Zenon Comp A 10μℓ, RT, more than 5min(less than a few days)

      2) Add Zenon Comp B 10μℓ to mixture 1(total 30μℓ), RT 5min

      3) Add PBT 70μℓ to mixture 2 (total 100μℓ)

Little trick

Reference